A cell is the minimum functional unit for exhibit life phenomenon. In an organism, cells having the same function assemble to constitute tissue, various types of tissues get together and cooperate to form an organ having a certain function, and the whole performs harmonious and unified vital activity. Many types of tissues are basically constituted by cells and extracellular matrices, and such a constitution is held by cells, cells adhering with extracellular matrix, extracellular matrix-extracellular matrix adhesion and cell-cell adhesion. It was thought that extracellular matrices have no bioactivity and are just fillers, but it has been becoming clear that they actually play an important role in almost all types of tissues. After cell-extracellular matrix adhesion activity was known, it was suggested that extracellular matrices not only play a role as a foothold for cells, but also are involved in the regulation of various cellular functions and the maintenance of homeostasis of tissues and organs. The importance of extracellular matrices has been more widely understood.
Cell-extracellular matrix adhesion is provided via a transmembrane cell-adhesion protein (typically integrin). Integrin is constituted by a heterodimer (α chain:β chain=1:1). 18 types of α chains and 8 types of β chains have been found, and at least 24 combinations thereof have been identified. It is known that each integrin recognizes a specific extracellular matrix (ligand). Moreover, it has been clarified that transmembrane cell-adhesion protein including integrin not only plays a role in cell-extracellular matrix adhesion/fixation, but also plays a role in converting information from extracellular matrices into intracellular signal to regulate proliferation, motility, death, differentiation, etc. of cells.
Integrins are classified into subfamilies based on specificity and function to ligand as follows: a collagen receptor; a laminin receptor; an RGD receptor recognizing an Arg-Gly-Asp (RGD) sequence contained in fibronectin, bitronectin or the like; and a leucocyte-specific receptor which is present only in leucocyte (Hynes R O. 2002. Integrins: Bidirectional, Allosteric Signaling Machines. Cell 110: 673-87; Miyasaka M. 2000. New edition of Adhesion Molecule handbook. Shujunsya). α4 and α9 integrins do not belong to the above-described subfamilies, and the subfamily thereof is referred to as α4 integrin subfamily (Elise L. Palmer, Curzio Rfiegg, Ronald Ferrando, Robert Pytela, Sheppard D. 1993. Sequence and Tissue Distribution of the Integrin α9 Subunit, a Novel Partner of 131 That Is Widely Distributed in Epithelia and Muscle. The Journal of Cell Biology 123: 1289-97).
Osteopontin (hereinafter abbreviated as OPN), which is one of extracellular matrices (ECM), is a secreted and acidic phosphorylated glycoprotein having the molecular weight of about 41 kDa. Expression of the molecule is widely recognized in breast fluid, urine, renal tubule, osteoclasts, osteoblasts, macrophages, activated T cells, tumor tissues, etc. The center portion of the molecule has: a cell-adhesive sequence GRGDS; and a SVVYGLR (SEQ ID NO:48) sequence (in human OPN) or a SLAYGLR (SEQ ID NO:47) sequence (in mouse OPN). Immediately after the position, there is a thrombin cleavage site. The molecule adheres to an integrin as an RGD receptor via the GRGDS sequence, and adheres to α4 (α4β1) and α9 (α9β1) integrins via the SVVYGLR (SEQ ID NO:48) sequence or the SLAYGLR sequence.
α4β1 binds to both a non-thrombin-cleaved OPN (non-cleaved OPN) and a thrombin-cleaved N-terminal fragment (cleaved OPN), while α9β1 only binds to a cleaved OPN. This difference of manner has already been found (Y Yokosaki et al., (1999) The Journal of Biological Chemistry 274, 36328-36334; P. M. Green et al., (2001) FEBS Letters 503, 75-79; S. T. Barry et al., (2000) Experimental Cell Research 258, 342-351). α4 and α9 integrins have many common ligands other than OPN. For example, an EDA site of fibronectin, a propeptide-von Willebrand factor (pp-vWF), tissue-type transglutaminase (tTG), blood coagulation factor XIII and Vascular Cell Adhesion Molecule-1 (VCAM-1) are known. Further, as a ligand specifically recognized by α4 integrin, CS-1 domain of fibronectin, MadCAM-1(α4β7) and the like are known. As a ligand specifically recognized by α9 integrin, tenascin C, plasmin and the like are known.
Amino acid sequences of α4 and α9 integrins and β1 integrin subunit are publicly known and are registered in GenBank. Further, it is known that amino acid sequences of these integrins have high similarity among species.
International Publication WO02/081522 discloses therapeutic effects on rheumatism-like arthritis and hepatitis by suppression of OPN function using an OPN-deficient mouse and a neutralization antibody to OPN. This publication also discloses that the SVVYGLR sequence, which is a recognition sequence of α4 integrin and α9 integrin, is important for onset of inflammatory diseases, and that a receptor to OPN is expressed in an immunocompetent cell and the like and is associated with inflammatory diseases.